Growth Assessment Of Multicellular Tumor Spheroids (MCTs)

Multicellular tumor (MCT) spheroids are an essential tool in tumor related research, and drug screens. MCT can be defined as aggregates of heterogeneous cells, closely packed with highly dense spheroids. In order to use MCTs in physiological research and clinical applications, it is important to characterize the nature, growth kinetics as well the response to chemo and radiation.

Evaluating the circularity of MCTs

The growth of MCTs is determined by its size and shape, which is primarily assessed through optical microscopy. 2D optical microscopic images are taken periodically and analyzed via software programs such as image J, to determine the circularity of spheroids. The circularity or “roundness” of the spheroids is in turn determined by the culturing method. A compact spheroid will typically increase in circularity over a few days. This increase is then followed by a decline in circularity over the remaining culture period indicating cell proliferation (1-2).

Assessing the size of MCTs

The size of MCTs is determined via measurement of two orthogonal diameters from optical microscope images. The size can be modulated by culture time, cell seeding density, and cell line type. It is necessary to accurately assess the spheroid size in order to produce reliable data that can be used for further study. A proper understanding of growth dynamics in parallel with the diverse factors that can affect growth is a necessary tool in standardizing MCTs cultures that are used for developing new treatment options as well as improving the existing treatment modalities (1-3).


1. Badea MA, Balas M, Hermenean A, Ciceu A, Herman H, Daniela I, et al. Influence of matrigel on single- and multiple-spheroid cultures in breast cancer research. SLAS Discov. 2019;24(5):563–78.

2. Raghavan S, Mehta P, Horst EN, Ward MR, Rowley KR, Mehta G. Comparative analysis of tumor spheroid generation techniques for differential in vitro drug toxicity. Oncotarget. 2016;7(13):16948–61.

3. Han, S.J., Kwon, S. & Kim, K.S. Challenges of applying multicellular tumor spheroids in preclinical phase. Cancer Cell Int 21, 152 (2021).

4. Mcmillan, Kay & Mccluskey, Anthony & Sorensen, Annette & Boyd, Marie & Zagnoni, Michele. (2015). Emulsion Technologies for Multicellular Tumour Spheroid Radiation Assays. The Analyst. 141. 10.